| 1. |
- Alizadehgharib, Sara, et al.
(författare)
-
Immunological response of human leucocytes after exposure to lipopolysaccharides from Porphyromonas gingivalis.
- 2021
-
Ingår i: Clinical and experimental dental research. - : Wiley. - 2057-4347. ; 7:4, s. 531-538
-
Tidskriftsartikel (refereegranskat)abstract
- Porphyromonas gingivalis (P. gingivalis) is a gram-negative bacterium and an important etiologic agent of periodontitis. P. gingivalis releases outer membrane vesicles containing lipopolysaccharides (LPS), which can penetrate periodontal tissues. Once in the periodontal tissues and in contact with immune cells, it may participate in the destructive innate host response associated with the disease. The exact mechanism of P. gingivalis LPS in the disease process is not clear, but it is known to affect a variety of immune responses.To investigate how LPS from P. gingivalis affect neutrophil extracellular trap (NET) formation, cell death and production of cytokines from human neutrophils and peripheral mononuclear blood mononuclear cells (PBMCs).Isolated neutrophils and PBMCs were cultured with LPS from P. gingivalis or Escherichia coli (E. coli) (control). The NET formation was measured using Sytox green stain. Cell death of neutrophils and PBMCs was analyzed using flow cytometry or Sytox green stain. Cytokine production was measured using enzyme-linked immunosorbent assay (ELISA) kit or Bio-Plex assay.Exposure to LPS from P. gingivalis and E. coli caused significantly lower cell death in neutrophils. NETs were formed after exposure to the two different LPS. In PBMCs, exposure to P. gingivalis and E. coli LPS caused increased levels of IL-1β and IL-6 compared to unstimulated controls. Increased cell death in PBMCs after exposure to LPS from E. coli in comparison to LPS from P. gingivalis and unstimulated controls was also observed.LPS from P. gingivalis has the ability to affect both human neutrophils and PBMCs with regard to cytokine production, cell death and production of NETs. LPS from P. gingivalis could be involved in the pathogenesis of periodontitis, and our results may contribute information regarding possible markers for diagnosis and targets for treatment of periodontal disease.
|
|
| 2. |
- Alizadehgharib, Sara, et al.
(författare)
-
The effects of the dental methacrylates TEGDMA, Bis-GMA, and UDMA on neutrophils in vitro.
- 2020
-
Ingår i: Clinical and experimental dental research. - : Wiley. - 2057-4347. ; 6:4, s. 439-447
-
Tidskriftsartikel (refereegranskat)abstract
- The prevalent usage of methacrylates in modern dentistry demands good knowledge of their biological impacts. While there have been several studies demonstrating the effects of different methacrylic monomers on mononuclear white blood cells, very little is known about the effects caused by these monomers on neutrophilic granulocytes. The objective of this study was to add novel knowledge about how neutrophils are affected by exposure to triethylene glycol dimethacrylate (TEGDMA), urethane dimethacrylate (UDMA), and bisphenol A glycol dimethacrylate (Bis-GMA) alone or in combinations.Isolated neutrophils were cultured in the presence or absence of methacrylates. The IL-8 release was measured using a DuoSet ELISA development kit. Apoptosis and necrosis were analyzed using flow cytometry. The formation of neutrophil extracellular traps (NETs) was investigated using Sytox green DNA staining combined with microscopically examination of released DNA and myeloperoxidase (MPO).The release of IL-8 was significantly increased after exposure to TEGDMA, Bis-GMA, UDMA, or TEGDMA in combination with Bis-GMA or UDMA compared to the unstimulated controls. Exposure to TEGDMA, UDMA, and Bis-GMA for 24hr separately or in combination did not affect apoptosis or necrosis of the exposed neutrophils. NET structures were formed by neutrophils after exposure to the different combinations of the methacrylates.The combination of TEGDMA and Bis-GMA had a synergistic proinflammatory effect on neutrophils by increasing the release of IL-8 and the formation of NET structures. The changes in the normal functions of neutrophils caused by methacrylate exposure may lead to altered inflammatory response and relate to previously reported adverse immune reactions caused by these substances.
|
|
| 3. |
- Björnsdottir, Halla, et al.
(författare)
-
Phenol-soluble Modulin α Peptide Toxins from aggressive Staphylococcus aureus induce rapid Formation of neutrophil extracellular Traps through a reactive Oxygen species-independent Pathway
- 2017
-
Ingår i: Frontiers in Immunology. - : Frontiers Media SA. - 1664-3224. ; 8
-
Tidskriftsartikel (refereegranskat)abstract
- Neutrophils have the ability to capture and kill microbes extracellularly through the formation of neutrophil extracellular traps (NETs). These are DNA and protein structures that neutrophils release extracellularly and are believed to function as a defense mechanism against microbes. The classic NET formation process, triggered by, e.g., bacteria, fungi, or by direct stimulation of protein kinase C through phorbol myristate acetate, is an active process that takes several hours and relies on the production of reactive oxygen species (ROS) that are further modified by myeloperoxidase (MPO). We show here that NET-like structures can also be formed by neutrophils after interaction with phenol-soluble modulin alpha (PSM alpha) that are cytotoxic membrane-disturbing peptides, secreted from community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA). The PSMa-induced NETs contained the typical protein markers and were able to capture microbes. The PSMa-induced NET structures were disintegrated upon prolonged exposure to DNase-positive S. aureus but not on exposure to DNase-negative Candida albicans. Opposed to classic NETosis, PSMa-triggered NET formation occurred very rapidly, independently of ROS or MPO, and was also manifest at 4 degrees C. These data indicate that rapid NETs release may result from cytotoxic membrane disturbance by PSMa peptides, a process that may be of importance for CA-MRSA virulence.
|
|
| 4. |
- Dahlstrand Rudin, Agnes
(författare)
-
Neutrophil recruitment in periodontal disease
- 2021
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Neutrophils are the first immune cells to arrive in infected or injured tissues, where they engulf microbes and clean up cell debris. Periodontitis is one of the typical symptoms of both neutropenia and defect neutrophil functionality, suggesting an important role for these cells in maintenance of periodontal health. While representing a minor fraction of the leukocytes in the periodontal lesion, neutrophils are the dominating cell type in the periodontal pocket and gingival crevicular fluid (GCF). The overall aim of this thesis was to characterize factors modulating neutrophil recruitment from blood to GCF in periodontitis. Neutrophil recruitment to the periodontal pocket is triggered by the bacterial species colonizing this site. Although previous studies have shown that subgingival bacteria trigger neutrophil chemotaxis, the bacterial chemoattractants responsible for this event remained to be identified. The aims of paper I and II were to identify soluble neutrophil chemoattractants released by the periodontitis associated bacterial species Porphyromonas gingivalis and Fusobacterium nucleatum, and their corresponding neutrophil receptors. Chemotactic compounds present in culture supernatants of both bacterial species where identified as short chain fatty acids (SCFAs) specifically activating neutrophils via the short chain fatty acid receptor 2 (FFAR2). CD177 is a neutrophil subtype marker with unknown function, expressed by 1–100% of circulating neutrophils depending on the donor. While CD177 has been proposed to facilitate neutrophil transmigration, this had not yet been demonstrated in vivo. The aim of paper III was to investigate whether CD177 expression affect neutrophil transmigration to GCF in periodontitis. The CD177+ subtype was enriched in GCF as compared to blood from the same donor, supporting an in vivo migration advantage of the CD177+ subtype to this site. Periodontitis patients also exhibited higher levels of CD177+ cells in blood as compared to healthy controls, which resulted in very high proportions of CD177+ cells in GCF. Considering this, functions differing between the subsets could influence destructive inflammation of the periodontal tissues. As CD177 may not be the sole factor contributing to functional differences between the subsets, further proteomic differences between CD177+ and CD177– neutrophils were investigated in paper IV. In conclusion, this thesis highlights SCFAs signaling via FFAR2 as factors involved in neutrophil chemotaxis triggered by periodontitis associated bacteria. Further, the CD177+ neutrophil subtype is preferentially recruited to GCF and functions specific for this subtype may be of importance for inducing (or suppressing) destructive inflammation in periodontal tissues.
|
|
| 5. |
- Dahlstrand Rudin, Agnes, et al.
(författare)
-
Porphyromonas gingivalis Produce Neutrophil Specific Chemoattractants Including Short Chain Fatty Acids
- 2021
-
Ingår i: Frontiers in cellular and infection microbiology. - : Frontiers Media SA. - 2235-2988. ; 10
-
Tidskriftsartikel (refereegranskat)abstract
- Neutrophil migration from blood to tissue-residing microbes is governed by a series of chemoattractant gradients of both endogenous and microbial origin. Periodontal disease is characterized by neutrophil accumulation in the gingival pocket, recruited by the subgingival biofilm consisting mainly of gram-negative, anaerobic and proteolytic species such as Porphyromonas gingivalis. The fact that neutrophils are the dominating cell type in the gingival pocket suggests that neutrophil-specific chemoattractants are released by subgingival bacteria, but characterization of chemoattractants released by subgingival biofilm species remains incomplete. In the present study we characterized small (< 3 kDa) soluble chemoattractants released by growing P. gingivalis, and show that these are selective for neutrophils. Most neutrophil chemoattractant receptors are expressed also by mononuclear phagocytes, the free fatty acid receptor 2 (FFAR2) being an exception. In agreement with the selective neutrophil recruitment, the chemotactic activity found in P. gingivalis supernatants was mediated in part by a mixture of short chain fatty acids (SCFAs) that are recognized by FFAR2, and other leukocytes (including monocytes) did not respond to SCFA stimulation. Although SCFAs, produced by bacterial fermentation of dietary fiber in the gut, has previously been shown to utilize FFAR2, our data demonstrate that the pronounced proteolytic metabolism employed by P. gingivalis (and likely also other subgingival biofilm bacteria associated with periodontal diseases) may result in the generation of SCFAs that attract neutrophils to the gingival pocket. This finding highlights the interaction between SCFAs and FFAR2 in the context of P. gingivalis colonization during periodontal disease, but may also have implications for other inflammatory pathologies involving proteolytic bacteria.
|
|
| 6. |
|
|
| 7. |
- Dahlstrand Rudin, Agnes, et al.
(författare)
-
Short chain fatty acids released by Fusobacterium nucleatum are neutrophil chemoattractants acting via free fatty acid receptor 2 (FFAR2)
- 2021
-
Ingår i: Cellular Microbiology. - : Hindawi Limited. - 1462-5814 .- 1462-5822. ; 23:8
-
Tidskriftsartikel (refereegranskat)abstract
- Fusobacterium nucleatum is a gram-negative and anaerobic oral commensal that is implicated in inflammatory conditions of the tooth-supporting structures, that is, periodontal diseases. One of the main characteristics of these conditions is an accumulation of neutrophil granulocytes in the gingival pockets where bacteria reside. Neutrophils are recruited to tissue-residing microbes by gradients of bacteria derived chemoattractants, and the cellular migration over the pocket epithelium into the gingival pocket is likely governed by chemoattractants released by the amino acid fermenting anaerobes typically colonising this site. However, the chemoattractants released by F. nucleatum and other oral anaerobes have long been unidentified. In the present study, we show that the major chemoattractants released during the growth of F. nucleatum are short chain fatty acids (SCFAs), primarily acetate and butyrate. These SCFAs, that are released at high levels as end-products of the metabolism of F. nucleatum, trigger chemotaxis of human neutrophils, as well as cytosolic Ca2+ signals, via free fatty acid receptor 2 (FFAR2). This finding establishes the SCFA-FFAR2 interaction as an important mechanism in the recruitment of neutrophils to the periodontal pocket, but could also be of importance in the pathogenesis of other medical conditions involving colonisation/infection of F. nucleatum.
|
|
| 8. |
- Dahlstrand Rudin, Agnes, et al.
(författare)
-
The neutrophil subset defined by CD177 expression is preferentially recruited to gingival crevicular fluid in periodontitis
- 2021
-
Ingår i: Journal of Leukocyte Biology. - : WILEY. - 0741-5400 .- 1938-3673. ; 109:2, s. 349-362
-
Tidskriftsartikel (refereegranskat)abstract
- In recent years, the concept of distinct subpopulations of human neutrophils has attracted much attention. One bona fide subset marker, exclusively expressed by a proportion of circulating neutrophils in a given individual, and therefore dividing neutrophils in two distinct subpopulations, is the glycoprotein CD177. CD177 is expressed on the plasma and granule membranes of 0-100% of circulating neutrophils depending on the donor. Several in vitro studies have linked CD177 to neutrophil transmigration, yet very few have looked at the role of CD177 for tissue recruitment in vivo. We investigate whether the CD177(+)and CD177(-)neutrophil subsets differ in their propensity to migrate to both aseptic- and microbe-triggered inflamed human tissues. Microbe-triggered neutrophil migration was evaluated in samples of gingival crevicular fluid (GCF) from patients with periodontitis, whereas neutrophil migration to aseptic inflammation was evaluated in synovial fluid from patients with inflammatory arthritis, as well as in exudate from experimental skin chambers applied on healthy donors. We found that the proportion of CD177(+)neutrophils was significantly higher in GCF from patients with periodontitis, as compared to blood from the same individuals. Such accumulation of CD177(+)neutrophils was not seen in the two models of aseptic inflammation. Moreover, the proportion of CD177(+)neutrophils in circulation was significantly higher in the periodontitis patient group, as compared to healthy donors. Our data indicate that the CD177(+)neutrophil subset is preferentially recruited to the gingival crevice of periodontitis patients, and may imply that this subtype is of particular importance for situations of microbe-driven inflammation.
|
|
| 9. |
- Davidsson, Lisa, et al.
(författare)
-
In Vivo Transmigrated Human Neutrophils Are Highly Primed for Intracellular Radical Production Induced by Monosodium Urate Crystals.
- 2020
-
Ingår i: International journal of molecular sciences. - : MDPI AG. - 1422-0067. ; 21:11
-
Tidskriftsartikel (refereegranskat)abstract
- Gout is an inflammatory disease caused by monosodium urate (MSU) crystals. The role of neutrophils in gout is less clear, although several studies have shown neutrophil extracellular trap (NET) formation in acutely inflamed joints of gout patients. MSU crystals are known to induce the production of reactive oxygen species (ROS) and NET formation in neutrophils isolated from blood, but there is inconclusive knowledge on the localization of ROS production as well as whether the ROS are required for NET formation. In this report we demonstrate that MSU crystals activate human neutrophils to produce ROS exclusively in intracellular compartments. Additionally, in vivo transmigrated neutrophils derived from experimental skin chambers displayed markedly increased ROS production as compared to resting blood neutrophils. We also confirmed that MSU stimulation potently induced NET formation, but this response was not primed in in vivo transmigrated neutrophils. In line with this we found that MSU-triggered NET formation was independent of ROS production and proceeded normally in neutrophils from patients with dysfunctional respiratory burst (chronic granulomatous disease (CGD) and complete myeloperoxidase (MPO) deficiency). Our data indicate that in vivo transmigrated neutrophils are markedly primed for oxidative responses to MSU crystals and that MSU triggered NET formation is independent of ROS production.
|
|
| 10. |
- Holdfeldt, André, et al.
(författare)
-
Reactivation of Gαi-coupled formyl peptide receptors is inhibited by Gαq-selective inhibitors when induced by signals generated by the platelet-activating factor receptor.
- 2017
-
Ingår i: Journal of Leukocyte Biology. - 0741-5400. ; 102:3, s. 871-880
-
Tidskriftsartikel (refereegranskat)abstract
- Formyl peptide receptor (FPR)-desensitized neutrophils display increased production/release of superoxide (O2(-)) when activated by platelet-activating factor (PAF), a priming of the response achieved through a unique receptor crosstalk mechanism. The aim of this study was to determine the effect of an inhibitor selective for small, heterotrimeric G proteins belonging to the Gαq subclass on that receptor crosstalk. We show that signals generated by FPRs and the PAF receptor (PAFR) induce activation of the neutrophil O2(-), producing NADPH-oxidase, and that response was sensitive to Gαq inhibition in cells activated by PAF, but no inhibition was obtained in cells activated by FPR agonists. Signaling in naive neutrophils is terminated fairly rapidly, and the receptors become homologously desensitized. The downstream sensitivity to Gαq inhibition in desensitized cells displaying increased production/release of O2(-) through the PAFR receptor crosstalk mechanism also comprised the reactivation of the FPRs, and the activation signals were redirected from the PAFR to the desensitized/reactivated FPRs. The Gαq-dependent activation signals generated by the PAFRs activate the Gαi-coupled FPRs, a receptor crosstalk that represents a novel pathway by which G protein-coupled receptors can be regulated and signaling can be turned on and off.
|
|
